Carriers of the apolipoprotein E (APOE) ?4 gene are at significantly increased risk for developing Alzheimer's disease (AD). Although numerous theories have been proposed, the cause of this association remains unclear. The most widely accepted view is that the accelerated AD pathology observed in APOE ?4 carriers is due to a decreased ability of the apoE4 protein to clear A? from the brain. However, possession of the APOE ?4 gene also results in a number of other neurological deficits unrelated to A? clearance, including alterations in neuronal structure, thinner entorhinal cortex (EC) layers and poorer outcomes after stroke, suggesting that there may be other mechanisms involved in this process. In order to gain a more comprehensive understanding of the how the expression of different apoE isoforms affects the brain and how this may impact the risk of developing AD and other age-related neurological illnesses, we propose to use mass spectrometry to identify lipids and small-molecules whose levels are affected by changes in apoE isoform expression. To accomplish this, we will first extract lipids, small-molecules and proteins from pathology-free EC and primary visual cortex (PVC) tissues obtained from 14-month old mice and postmortem 19-55 year old individuals expressing differing apoE isoforms. The lipid and small-molecule fractions from these extracts will then be used to perform targeted lipidomics and untargeted metabolomics, followed by bioinformatic analysis and a variety of validation experiments, in order to determine the specific lipids, small-molecules and metabolic pathways that are affected by alternative apoE isoform expression in the brain. Thus far, preliminary studies have uncovered significant changes in energy metabolism pathways and several important lipid subclasses, demonstrating the power of these techniques for discovering previously unknown isoform-specific effects of apoE in the brain. We expect that the full study proposed herein will uncover further apoE isoform-specific changes in lipid and small-molecule levels and will lead to a greater understanding of how apoE4 influences AD pathology, potentially leading to new therapeutic strategies for AD and other neurological illnesses influenced by apoE4 expression.